Medicament for treatment of non-insulin dependent diabetes mellitus, hypertension and/or the metabolic syndrome

ABSTRACT

A substance including the chemical structures of bicyclo [3.2.1]octan or the chemical structures of kaurene for the use in a dietary supplementation or as a constituent in a medicament for the treatment of non-insulin dependent diabetes mellitus, hypertension and/or the metabolic syndrome. The unique chemical structures of bicyclo [3.2.1]octan alone or in a kaurene structure provides the substances, such as e.g. steviol, isosteviol and stevioside with the capability of enhancing or potentiating the secretion of insulin in a plasma glucose dependent manner. The substances including these unique chemical structures also have the capability of reducing the glucagon concentration in the blood and/or lowering the blood pressure thereby providing a self-regulatory treatment system for non-insulin dependent diabetes mellitus and/or hypertension. In a combination drug which also comprise a soy protein, and/or soy fiber and/or at least one isoflavone these substances act synergistically and such combination drugs are highly useful both prophylacticly or directly in the treatment of e.g. the metabolic syndrome and obesity and has due to the self-regulatory effect a widespread applicability as a dietary supplementation.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a division of U.S. application Ser. No. 10/210,787filed Jul. 31, 2002, which is a continuation of Internationalapplication PCT/DK01/00075 filed Feb. 1, 2001, the entire content ofeach of which is expressly incorporated herein by reference thereto.

TECHNICAL FIELD

The present invention relates to a new medicament for the treatment ofnon-insulin dependent diabetes mellitus, hypertension, metabolicsyndromes and other conditions in mammals.

BACKGROUND ART

Diabetes is a common disease that has a prevalence of 2-4% in thepopulation. Non-insulin dependent diabetes mellitus comprises about 85%of diabetes most commonly occurring at the age above 40 years. Theincidence of non-insulin dependent diabetes mellitus is increasing andis at a global level expected to surpass 200 million subjects at year2010.

Diabetes is associated with increased morbidity and a 2-4-fold increasein mortality primarily due to cardiovascular diseases and strokes.

Non-insulin dependent diabetes mellitus develops especially in subjectswith insulin resistance and a cluster of cardiovascular risk factorssuch as obesity, hypertension and dyslipidemia, a syndrome which firstrecently has been recognized and is named “The metabolic syndrome”(Alberti K. G., Zimmet P. Z.; Definition, diagnosis and classificationof diabetes mellitus and its complications”. Part 1: Diagnosis andclassification of diabetes mellitus provisional report of a WHOconsultation. Diabet. Med. July 1998; 15(7), p. 539-53).

In accordance with the WHO-definition (www.idi.org.au/whoreport.htm), apatient has metabolic syndrome if insulin resistance and/or glucoseintolerance is present together with two or more of the followingconditions:

-   -   reduced glucose tolerance or diabetes    -   insulin sensivity (under hyperinsulinaemic, euglycaemic        conditions corresponding to a glucose uptake below the lower        quartile for the background population)    -   increased blood pressure (≧140/90 mmHg)    -   increased plasma triglyceride (≧1.7 mmol/l) and/or low HDL        cholesterol (<0.9 mmol/l for men; <1.0 mmol/l for women)    -   central adipositas (waist/hip ratio for men: >0.90 and for        women >0.85) and/or Body Mass Index >30 kg/m²)    -   micro albuminuria (urine albumin excretion: >20 μg min⁻¹ or        albumin/creatinine ratio ≧2.0 mg/mmol.

It has become increasingly evident that the treatment should aim atsimultaneously normalizing blood glucose, blood pressure, lipids andbody weight to reduce the morbidity and mortality. Diet treatment,exercise and avoiding smoking are the first treatment modalities thatshould be started. However, it will often be necessary to addpharmacological therapy but until today no single drug thatsimultaneously attacks hyperglycaemia, hypertension and dyslipidemia areavailable for patients with metabolic syndrome. Instead, these patientsmay be treated with a combination of several different drugs in additionto e.g., diet. This type of treatment is difficult to adjust andadminister to the patient and such treatment may result in many unwantedadverse effects which in themselves may need medical treatment.

Consequently there is a long felt need for a new and combined medicamentfor the treatment of metabolic syndrome thereby also preventing anincrease in the number of persons developing the non-insulin dependentdiabetes mellitus.

Existing oral antidiabetic medicaments to be used in such treatmentinclude the classic insulinotropic agents sulphonylureas (Lebovitz H. E.1997. “The oral hypoglycemic agents”. In: Ellenberg and Rifkin'sDiabetes Mellitus. D. J. Porte and R. S. Sherwin, Editors: Appleton andLange, p. 761-788). They act primarily by stimulating thesulphonylurea-receptor on the insulin producing beta-cells via closureof the K⁺ _(ATP)-sensitive channels. However if such an action alsoaffects the myocytes in the heart, an increased risk of cardiacarrhytmias might be present. Also, it is well know in the art thatsulphonylureas can cause severe and lifethreatening hypoglycemia, due totheir continuous action as long as they are present in the blood.

Consumption of soy protein rather than animal protein has been found tolower blood cholesterol (Anderson J. W., Johnstone B. M., Cook-Newell M.E.: Meta-analysis of the effects of soy protein intake on serum lipids.N. Engl. J. Med. 1995; 333; p. 276-282). In addition to this knowledge,recent research also provides evidence that soy protein and/orisoflavones may improve endothelial function and attenuate eventsleading to both lesion and thrombus formation (Anderson J. W., JohnstoneB. M., Cook-Newell M. E.: “Meta-analysis of the effects of soy proteinintake on serum lipids”; N. Engl. J. Med. 1995; 333; p. 276-282; PotterS. M., Soy protein and cardiovascular disease: “The impact of bioactivecomponents in soy”. Nutrition Reviews 1998;56, p. 231-235).

Several attempts to develop new antidiabetic agents and drugs for thetreatment or prophylactic treatment of the syndrome not having theadverse effects mentioned above, e.g. hypoglycemia and potential harmfulactions on the heart functions have been made over the years. For thispurpose, plants provide a vast resource of compounds with the potentialto become new antidiabetic agents.

For instance extracts of the leaves of Stevia rebaudiana Bertoni, aherbaceous member of the Compositae family, have been used for manyyears in the treatment of diabetes among Indians in Paraguay and Brazil(Sakaguschi M., Kan P Aspesquisas japonesas com Stevia rebaudiana (Bert)Bertoni e o estevioside. Cienc. Cultur. 34; p. 235-248,1982; Oviedo C.A., Franciani G., Moreno R., et al. “Action hipoglucemiante de la SteviaRebaudiana Bertoni (Kaa-he-e)”. Excerpt. Med. 209, p. 92,1979; Curi R.,Alvarez M., Bazotte R. B., et al. Effect of Stevia rebaudiana on glucosetolerance in normal adult humans. Braz. J. Med. Biol. Res., 19, p.771-774, 1986; Hansson J. R., Oliveira B. H., “Stevioside and relatedsweet diterpenoid glycoside”. Nat. Prod. Rep. 21, p.301-309, 1993).

Also, an antihyperglycemic effect has been found in rats whensupplementing the diet with dried S. rebaudiana leaves (Oviedo C. A.,Franciani G., Moreno R., et al. “Action hipoglucemiante de la SteviaRebaudiana Bertoni (Kaa-he-e)”. Excerpt. Med. 209:92, 1979). Curi et al.found a slight suppression of plasma glucose when extracts of Steviarebaudiana leaves were taken orally during a 3-day period. Furthermore,Oviedo et al. reported that tea prepared from the leaves caused a 35%reduction in blood glucose in man.

A number of Stevia species have been examined and shown to containlabdanes, clerodanes, kaurenes and beyerenes (Hansson J. R., Oliveira B.H., “Stevioside and related sweet diterpenoid glycoside”. Nat. Prod.Rep. 21, p. 301-309, 1993). Any of these substances as well as manyothers unidentified substances in the leaves could be responsible forthe reduction in blood glucose in man.

In the work of Malaisse W.J. et al (Malaisse W. J., Vanonderbergen A.,Louchami K, Jijakli H. and Malaisse-Lagae F., “Effects of ArtificialSweeteners on Insulin Release and Cationic Fluxes in Rat PancreaticIslets”, Cell. Signal. Vol 10, No. 10, p. 727-733, 1998) the effect ofseveral artificial sweeteners, including stevioside, on insulin releasefrom isolated normal pancreatic rat islets were studied. In this studyit was reported that in the presence of 7 mmol/l D-glucose, steviosidein a concentration of 1.0 mmol/l caused a significant increase ininsulin output. Also the control group demonstrated a significantincrease in insulin output of about 16 times above the basal releasevalue in the presence of 20 mmol/l D-glucose increase. It is thereforeuncertain whether the insulin releasing effect is due to the increasedglucose level or the presence of stevioside. No diabetic islet cellswere studied and the skilled person within the art will know that themechanism for stimulating normal pancreatic islet cells either notfunctions at its optimum or not functions at all in the diabeticpancreatic cells, and that the study provided no certain indication ofthe possible use of stevioside in the treatment of non-insulin dependentdiabetes mellitus, hypertension and/or the metabolic syndrome.

In a Chinese study (Lin Qi-Xian, Cao Hai-Xing, Xie Dong, Li Xing-Ming,Shang Ting-Lan, Chen Ya-Sen, Ju Rui-Fen, Dong Li-Li, Wang Ye-Wen, QuianBao-Gong, “Experiment of Extraction of Stevioside”, Chinese Journal ogPharmaceuticals 1991, No. 22, p 389-390) is indicated a method forextracting stevioside from stevioside leafs from the origin of Bingzzhouin the Hunan Province. The content of stevioside in the extract wasdetermined using HPLC although the article is silent of the purity ofthe extract. The produced stevioside tablets were for no apparent reasonand medical indication applied to patients in the Wuhan Second Hospital.No data on the influence of stevioside on blood glucose, insulin and/orblood pressure is revealed. It is stated that the tablets were effectiveto diabetes and hypertension during preliminary clinical observations.However, total lack of data on blood glucose, insulin and/or bloodpressure i.e., lack of support by test results and the missinginformation of which types of diabetes that were treated, makes this anunsupported and unconfirmed assertion.

Any detailed information of which substance or substances in the leavesthat might cause a possible anti-hyperglycemic effect has not yet beendisclosed for certainty, and the mechanism of how and to which extentthe plasma glucose is reduced is unknown. The above mentioned articlesand studies are concerned with the initial discovery of the effects andprovide no evidence of which specific component(s) in the leaves thatmight be the active one(s).

The effect of intravenous stevioside on the blood pressure was studiedin spontaneously hypertensive rats (“The Effect of Stevioside on BloodPressure and Plasma Catecholamines in Spontaneously Hypertensive Rats”,Paul Chan, De-Yi Xu, Ju-Chi Liu, Yi-Jen Chen, Brian Tomlinson, Wen-PinHuang, Juei-Tang Cheng, Life Science, Vol. 63, No. 19, 1998, p.1679-1684). The study showed that during an intravenously administrationof stevioside of 200 mg/kg the hypotensive effect was at a maximum, butalthough reported as being significantly the fall in the systolic bloodpressure was only small. Neither the heart rate nor the plasmacatecholamines were significantly changed during the observation period.This study indicated that stevioside advantageously could be used fortreating hypertension.

No reports of an effect on plasma glucagon level have previously beenreported. Glucagon, a pancreatic islet hormone, acts as a diabetogenichormone by increasing the hepatic glucose output thereby elevating bloodglucose.

Recent studies and tests made by the present inventors have focused onespecially the diterpenoid glycoside stevioside which is a majorconstituent found in the leaves of Stevia rebaudiana where it may occurin amounts of up to about 10% (Hansson J. R., Oliveira B. H.,“Stevioside and related sweet diterpenoid glycoside”. Nat. Prod. Rep.21, p.301-309, 1993; Bridel M., Lavielle R., Physiologie Vegetale: “Surle principe sucre'du Kaa' he'e (Stevia rebaudiana Bertoni): II Lesproduits d'hydrolyse diastasique du stevioside, glucose et steviol”.Acad. Sci. Paris 192, p. 1123-1125, 1931; Soejarto D. D., Kinghorn A.D., Farnsworth N. R., Potential sweetening agent of plant origin. III:“Organoleptic evaluation of Stevia leaf herbarium samples forsweetness”. J. Nat. Prod. 45, p. 590-598, 1983; Mossettig E., Nes W. E.Stevioside. II: “The structure of the aglucone”; J. Org. Chem. 20, p.884-899, 1955; Kohda H., Hasai R., Yamasaki K. et al. “New sweetditerpene glucosides from Stevia rebaudiana”. Phytochemistry 15, p.981-983, 1976).

Also, its aglycone, steviol, has been found to be contained in theleaves of Stevia rebaudiana as well as other sweet-tasting glycosidese.g. Steviolbioside, Rebaudioside A,B,C,D and E, and Dulcoside (BridelM., Lavielle R., Physiologie Vegetale: “Sur le principe sucre'du Kaa'he'e (Stevia rebaudiana Bertoni): II Les produits d'hydrolysediastasique du stevioside, glucose et steviol”. Acad. Sci. Paris 192, p.1123-1125, 1931; Soejarto D. D., Kinghorn A. D., Farnsworth N. R.,Potential sweetening agent of plant origin. III: “Organolepticevaluation of Stevia leaf herbarium samples for sweetness”. J. Nat.Prod. 45, p. 590-598, 1983; Mossettig E., Nes W. E. Stevioside. II: “Thestructure of the aglucone”; J. Org. Chem. 20, p. 884-899, 1955;Mossettig E., Nes W. E. Stevioside. II: “The structure of the aglucone”;J. Org. Chem. 20, p. 884-899, 1955; Kohda H., Hasai R., Yamasaki K. etal. “New sweet diterpene glucosides from Stevia rebaudiana”.Phytochemistry 15, p. 981-983, 1976).

The present inventors have already successfully proved that bothstevioside and steviol have an anti-hyperglycemic, glucagonostatic andinsulinotropic effect when administered intravenously to rats and astimulatory effect on the insulin secretion from mouse islets in vitro.

No well defined, chemical stable, non-toxic, reliable and non-adverseeffects alternative to the sulphonylureas for the treatment ofnon-insulin dependent diabetes mellitus is available today, however, andthese findings have given rise to further studies and tests of analoguesand derivates of these substances in order to find improved andalternative drugs for a self-regulatory treatment of diabetes,hypertension and especially metabolic syndrome in mammals, andpreferably in humans.

In order to prevent sequelae or to delay the developing of a number ofthe above-mentioned metabolic and functional disorders in humans, thereis a need it for new and beneficial dietary supplementations or newself-administrable non-prescription drugs for prophylaxis. The presentinvention now satisfies this need.

SUMMARY OF THE INVENTION

Accordingly, the present invention relates to a selectively responsivemedicament composition comprising at least one substance including abicyclo [3.2.1]octan in a double ring system having a basic chemicalskeletal of a kaurene structure having the structural formula II:

or an analogue, derivative or metabolite thereof, wherein the substanceresponds only at an elevated plasma glucose concentrations. Generally,the response of the substance is initiated by a plasma glucoseconcentration of 6 mmol/l or larger.

Preferably, the substance is selected from the group consisting ofsteviol, isosteviol, glucosilsteviol, gymnemic acid, steviolbioside,stevioside Rebaudioside A, Rebaudioside B, Rebaudioside C, RebaudiosideD, Rebaudioside E and Dulcoside A, their pharmaceutically acceptableanalogues or their pharmaceutically acceptable derivates. The substancecan be isolated from a plant source and can be used alone or incombination with at least one soy protein alone or in combination withat least one isoflavone.

The substance and composition can be used as a dietary supplement or asa medicament for a mammal. As noted above the substance or compositionis responsive in the mammal only when the mammal's plasma glucoseconcentrations are elevated. Thus, the medicament can be used fortreating the mammal for non-insulin dependent diabetes mellitus,metabolic syndrome, to stimulate insulin production, to reduce glucagonconcentrations, to suppress fasting plasma triglycerides or totalcholesterol levels in the mammal, or for treating hypertension in themammal. Preferably, the medicament is an oral medicament and isself-regulating.

The invention also relates to a method of making a selectivelyresponsive composition which comprises associating with a carrier abicyclo [3.2.1]octan in a double ring system having a basic chemicalskeletal of a kaurene structure having the structural formula II,wherein the substance responds only at an elevated plasma glucoseconcentrations. The composition that is made can be used as a dietarysupplement or as one of the medicaments mentioned above.

The invention also relates to various treatment methods for mammals,including treating non-insulin dependent diabetes mellitus, treatingmetabolic syndrome, treating hypertension, suppressing fasting plasmatriglycerides, suppressing total cholesterol level, or suppressingappetite.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention is further illustrated by the following examples and theaccompanying drawings that are intended to illustrate preferred featuresand properties of the invention, wherein:

FIG. 1 shows the chemical structure of steviol, isosteviol andstevioside,

FIG. 2 a shows the effect of stevioside on blood glucose during i.v.glucose tolerance test in normal Wistar rats,

FIG. 2 b shows the effect of stevioside on blood glucose during i.v.glucose tolerance test in GK rats,

FIG. 3 a shows the effect of stevioside on glucose-induced releaseduring i.v. glucose tolerance test in normal Wistar rats,

FIG. 3 b shows the effect of stevioside on glucose-induced releaseduring i.v. glucose tolerance test in GK rats,

FIG. 4 a shows the effect of stevioside on glucose-stimulated insulinsecretion from isolated mouse islets,

FIG. 4 b shows the effect of steviol on glucose-stimulated insulinsecretion from isolated mouse islets,

FIG. 5 a shows the effect of an i.v. bolus injection of glucose onplasma glucagon levels during an intravenous glucose tolerance test inGK rats,

FIG. 5 b shows the effect of an i.v. bolus injection of glucose andstevioside on plasma glucagon levels during a glucose tolerance test inGK rats,

FIG. 6 a shows the systolic blood pressure during 6 weeks treatment ofGK rats with stevioside,

FIG. 6 b shows the diastolic blood pressure in GK rats treated withstevioside.

FIG. 7 a shows the effect of 10⁻³ mmol/l stevioside on the insulinsecretion from isolated mouse islets in the presence of glucose rangingbetween 0 and 16.7 mmol/l,

FIG. 7 b shows the effect of 10⁻⁶ mmol/l steviol on the insulinsecretion from isolated mouse islets in the presence of glucose rangingbetween 0 and 16.7 mmol/l,

FIG. 8 a-d shows the acute effects of stevioside in type II diabeticpatients, and

FIG. 9 a-g shows the effects of the action of the combination ofstevioside and soy based dietary supplementation in diabetic GK-rats.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

Careful structural chemistry studies by the inventors have revealed thatall potential substances for stimulating the insulin secretion extractedfrom the leaves of Stevia rebaudiana share the common unique skeletalstructure of bicyclo[3.2.1]octan of the formula I:

This bicyclo[3.2.1]octan can be found in e.g. steviol, isosteviol and instevioside. The formula I structure has also been recognised inglucosilsteviol, gymnemic acid, steviolbioside, Rebaudioside A,Rebaudioside B, Rebaudioside C, Rebaudioside D, Rebaudioside E andDulcoside A.

All these substances also share the common structure of formula II:

which is the basic structure in kaur-16-en-18-oic acid.

These specific structures of the formula I or II are recognized inseveral chemical compounds, which have been shown to have a highlypotent insulin stimulating effect on isolated mouse pancreatic β-cell,and these structures of formula I and II are evidently the active partsof the molecules in causing the stimulating task.

This assumption is further confirmed by the fact that tests have shownthat steviol having the smallest skeletal structure stimulate theinsulin secretion to a greater extent than e.g. the glycoside steviosidehaving a much larger skeletal structure. Also, the inventors of thepresent invention have succeeded in purifying the differentRebaudiosides from Stevia rebaudiana and preclinical animal studiesindicate the same stimulatory effect on insulin secretion.

Consequently this indicates that other compounds including thestructures of the formula I or II, such as e.g. analogues, derivates andmetabolites of the compounds mentioned above, can be used alternatively.

Studies and tests on rats have disclosed that the insulin stimulatingeffect of these substances is dependent on the concentration of theplasma glucose.

The substances comprising the chemical structures, which includes theformula I or II, did not cause an insulin release as long as the plasmaglucose concentration was below approximately 6 mmol/l. At plasmaglucose concentration above 6 mmol/l, the stimulating effect of thecompounds provided an elevated plasma insulin concentration resulting inan immediate suppression of plasma glucose concentration thereby keepingthis at a normal level.

In addition to the above findings, the present inventors havesurprisingly found that the substances comprising the chemicalstructures including the formula I or II also have the capabilities ofreducing the glucagon concentration in the blood.

This characteristic nature and qualities of the substances make them anobvious choice as a component in a medicament for the treatment ofespecially non-insulin dependent diabetes mellitus (NIDDM).

The finding that e.g. intravenously administered stevioside inhibitedblood glucose responses to intravenous glucose in NIDDM rats (GK rats)but not in normal rats supports this fact. This finding is new andsurprisingly has neither been expected nor demonstrated in earlierstudies that have only been concerned with normal pancreatic isletcells.

As a further example of the unique action of the substances according tothe invention, stevioside infusion at normal blood glucose did not causeany hypoglycemia irrespective of it being given as a bolus or at aconstant intravenous infusion.

Due to the insulin secretory stimulating effect induced by a slightlyelevated plasma glucose concentration, the simultaneous plasma glucagonreducing effect and the inhibited blood glucose response, thesesubstances are able to control, regulate and adjust the plasma glucoseconcentration of a NIDDM patient to a normal level.

As a consequence of the glucose-dependency the substances only act whenneeded, e.g. after the patient has increased blood glucose after havingeaten. In NIDDM patients treated with medicaments including thesesubstances hypoglycemia will not occur and hypoglycemia will becounteracted.

Therefore, the substances provide a self-regulatory system respondingonly at elevated plasma glucose concentration.

The substances are preferably used in medicaments for oral medication.When taken orally, the glycosylated substances can be partiallymetabolised but the basic skeletal structure of the formula I or II willnot be changed and the different characteristic effects mentioned abovewill be preserved.

The treatment with a medicament including these substances provides anattractive alternative to different types of drugs available andpresently used today for the treatment of NIDDM, such drugs being drugsfor stimulating the insulin secretion (sulphonylureas or repaglinide),drugs for improving the insulin sensivity (biguanides andthiazolidinediones) or drugs for retarding gastrointestinal carbohydrateabsorption (α-glucosidase inhibitors).

The potential of these new substances has for the first time also beentested in human NIDDM studies and the beneficial and advantageouslycombined multiple effects in humans of a single substance according tothe invention has been demonstrated and will be further described in theexamples.

The above-mentioned human tests have been conducted by orallyadministrating the substances, but within the scope of the invention thesubstances can optionally be used in the preparation of medicaments forintravenous, subcutaneous or intramuscular medication.

The substances further bring along the blood pressure reducing effect.In long-term experiments stevioside acutely suppresses blood pressure indiabetic rat. This important discovery is of the benefit to the diabeticpatients that have developed hypertension in relation to or besidestheir disease.

When at least one of the substances according to the invention iscombined in a medicament also comprising at least one soy protein aloneor in combination with at least one isoflavone, it is possible tomanufacture a combined preparation of a drug for the treatment ofpatients with the metabolic syndrome in accordance with the previouslydefinition. Such a medicament may advantageously be used in prophylactictreatment of patient in a risk group. For example, a slow-release drugon the basis composition mentioned above provides a convenient treatmentfor the patient with the metabolic syndrome.

The inventors of the present invention have demonstrated that thecombination of the substances according to the invention and at leastone soy protein have a new unexpected and surprisingly synergisticeffect surpassing the additive effect of the single components of themedicament thereby providing a completely new and very importantmedicament for therapeutic or prophylactic treatment of the metabolicsyndrome.

The present inventors have used the combination of the substancesaccording to the invention and at least one soy protein as a dietarysupplementation in human studies. The test results significantly proved,as will be seen in the following examples, that such combination has abeneficial impact on cardiovascular risk markers in type II diabeticsubjects.

Stevioside at a dose as high as 15 g/kg body weight was not lethal toeither mice, rats or hamsters (Toskulkao C., Chaturat L., Temcharoen P.,Glinsukon T. “Acute toxicity of stevioside, a natural sweetener, and itsmetabolite, steviol, in several animal species”. Drug Chem. Toxicol.February-May 1997; 20(1-2), p. 31-44). In rats and mice, LD₅₀ values ofsteviol were higher than 15 g/kg body weight while the LD₅₀ for hamsterswere 5-6 g/kg body weight. The latter was accompanied with degenerationof the proximal tubular cells, which correlated to increases in bloodurea nitrogen and creatinine. Stevioside is excreted by the urine (MelisM. S. “Renal excretion of stevioside in rats”. J. Nat. Prod. May 1992;55(5), p. 688-90) and is not metabolised in the isolated perfused ratliver (Ishii-Iwamoto E. L., Bracht A. “Stevioside is not metabolised inthe isolated perfused rat liver”. Res. Commun. Mol. Pathol. Pharmacol.February 1995; 87(2), p. 167-75).

Stevioside and steviol showed no mutagenic effect on a number ofSalmonella typhimurium strains (Klongpanichpak S., Temcharoen P.,Toskulkao C., Apibal S., Glinsukon T. “Lack of mutagenicity ofstevioside and steviol in Salmonella typhimurium TA 98 and TA 100”. J.Med. Assoc. Thai September 1997; 80 Suppl. 1, p. 121-128; Suttajit M.,Vinitketkaumnuen U., Meevatee U., Buddhasukh D. “Mutagenicity and humanchromosomal effect of stevioside, a sweetener from Stevia rebaudianaBertoni”. Environ Health Perspect October 1993; 101 Suppl. 3, p. 53-56).In another study, it was confirmed that stevioside was not mutagenicwhereas steviol, however, produced dose-related positive responses insome mutagenicity test (Matsui M., Matsui K., Kawasaki Y., Oda Y.,Noguchi T., Kitagawa Y., Sawada M., Hayashi M., Nohmi T., Yoshihira K.,Ishidate M. Jr., Sofuni T. “Evaluation of the genotoxicity of steviosideand steviol using six in vitro and one in vivo mutagenicity assays”.Mutagenesis November 1996; 11(6), p. 573-579).

Stevioside is not carcinogenic in F344 rats (Toyoda K., Matsui H., ShodaT., Uneyama C., Takada K., Takahashi M. “Assessment of thecarcinogenicity of stevioside in F344 rats”. Food Chem. Toxicol. June1997; 35(6), p. 597-603). Doses as high as 2.5 g/kg body weight/day hadno effect on growth or reproduction in hamsters (Yodyingyuad V.,Bunyawong S. “Effect of stevioside on growth and reproduction”. Hum.Reprod. January 1991; 6(1), p. 158-165).

To the knowledge of the inventors, no observations or reports showingpotential toxic effects in humans have been published.

It will be recognized by the skilled artisan that rearranged structuresof the formula II are within the scope of the invention, and suchrearrangements might occur naturally in the gastro intestinal tract. Asexample can be mentioned that rearrangement may occur at the C16 forminga double bond to the C15 and thereby leaving a single bond open forsubstitution at position 17. A COOH group at position 18 is open for anumber of reactions such as reaction with alcohol, as well as a numberof substituents can be provided at any point of the formula IIstructure. Also, other substituents such as e.g. saccharides, at thevarious C-atoms and the structures may be anticipated.

EXAMPLES

In the following examples, the type II diabetic Goto-Kakizaki (GK) ratsoriginated from Takeda Chemical Ind., Tokyo, Japan and were bredlocally.

The normal Wistar rats and the NMRI mice were available fromBomholtg{dot over (a)}rd Breeding and Research Centre Ltd., Ry, Denmark.

The rats had a weight of 300-350 g and the mice a weight of 22-25 g. Theanimals were kept on a standard pellet diet and tap water ad libi tum.

The stevioside is obtained from the Japanese company WAKO-TriCHEM.

The abbreviation IAUC means Incremental Area Under the Curve (abovebasal).

Example 1

As examples of the effects of a compound including the chemical formulasII, stevioside was tested on normal Wistar rats and on GK rats. 2.0 gglucose/kg body weight and 0.2 g stevioside/kg body weight weredissolved in 0.9% saline and infused intravenously. The plasma glucoseand insulin levels were measured over a period of 2 hours.

The results are shown in FIGS. 2 a, 2 b, 3 a and 3 b, were the O-Oseries (n=6 for Wistar and n=14 for GK) illustrate glucose infused aloneand the {circle over (2)}-{circle over (2)} series (n=6 for Wistar andn=12 for GK) illustrate the combined glucose and stevioside infusion.Data are given as mean±SEM.

After administration of the glucose load, plasma glucose raisedimmediately and plasma insulin raised abruptly. When stevioside wasadded together with the glucose, a diminished glucose response was foundin the GK-rat and a significant decrease was observed already after 30min. In the GK rat, stevioside caused a pronounced increase in theinsulin response compared to the Wistar rat. The stevioside-inducedinsulin response was delayed and increased throughout the whole test.The insulin response was monophasic.

This discovery of stevioside having a blood glucose reducing effect inthe type II diabetic rat indicates that stevioside and compounds havinga similar chemical structure can be used in a medicament for thetreatment of NIDDM in man.

Example 2

Islet from 6-10 NMRI mice were isolated and incubated in the presence of16.7 mmol/l and 10⁻⁹-10⁻³ mol/l stevioside or 10⁻⁹-10⁻³ mol/l steviol.

The results of these tests are illustrated in FIGS. 4 a and 4 b whereeach column represents mean±SEM from 24 incubations of single islets.Black bars in FIG. 4 a indicate that stevioside is present and hatchedbars indicate that stevioside is absent.

Black bars in FIG. 4 b indicate that steviol is present and hatched barsindicate that steviol is absent.

The figures show that stevioside and steviol are capable of potentiatingglucose-stimulated insulin secretion. Further tests confirmed that astimulatory effect was found already at a very low concentration (above0.1 nM).

Example 3

During a glucose tolerance test, an intravenous bolus of stevioside of0.2 g/kg body weight was injected in GK rats (the {circle over(2)}-{circle over (2)} serie (n=6)). GK rats receiving 0.9% salineintravenously served as controls (the O-O serie (n=6)). Glucose 2.0 g/kgbody weight was administered as a bolus at timepoint 0 min. The plasmaglucagon responses are shown as mean±SEM in FIGS. 5 a (control) and 5 b(GK). The plasma glucagon was suppressed in the stevioside treated GKrat.

Example 4

GK rats were treated with stevioside 0.025 g/kg body weight/24 h for 6weeks. Stevioside was administered in the drinking water. GK ratsreceiving drinking water with 0.111 g D-glucose/kg body weight/24 hserved as controls. Systolic (FIG. 6 a, control: O-O series,stevioside-treated: {circle over (2)}-{circle over (2)} series) anddiastolic (FIG. 6 b, control: O-O series, stevioside-treated: {circleover (2)}-{circle over (2)} series) blood pressures were measured on thetail.

The figures show a 10-15% decrease in the blood pressure detectableafter 2 weeks of treatment and the effect hereafter was stable andconsistent during the study period.

Example 5

The influence of the maximal stimulatory doses of 10⁻³ mol/l steviosideand 10⁻⁶ mol/l steviol was studied in NMRI mouse islets over a rangebetween 0 and 16.7 mmol/l glucose. Both stevioside (FIG. 7 a) andsteviol (FIG. 7 b) potentiated insulin secretion at and above 8.3 mmol/land indicated that the initiating level for stimulating insulinsecretion was between 3.3 mmol/l and 8.3 mmol/l of glucose. Black barsin FIG. 7 a indicate that stevioside is present and hatched barsindicate that stevioside is absent. Black bars in FIG. 7 b indicate thatsteviol is present and hatched bars indicate that steviol is absent.

Example 6

Twenty type II diabetic patients (6 female/14 males) with a mean age of63.6±7.5 years participated in a controlled randomised double blindcrossover trial. They were supplemented for 6 weeks with soy protein for(50 g/day) with high levels of isoflavones (minimum 165 mg/day) andcotyledon fibers (20 g/day) or placebo (casein 50 g/day) and cellulose(20 g/day) separated by a 3 week wash-out period.

This dietary supplement significantly reduced LDL-Cholesterol by 10%(p<0.05), LDL/HDL ratio by 12% (p<0.05), Apo B-100 by 30% (p<0.01),triglycerides by 22% (p<0.05) and homocystein by 14% (p<0.01). No changewas observed in HDL-Cholesterol, Factor VIIc, von Willebrandt factor,fibrinogen, PAI-1, HbAlc or 24 hour blood pressure.

The results indicate beneficial effects of dietary supplementation withsoy protein on cardiovascular risk markers in type II diabetic subjects.The improvement is also seen in individuals with near-normal lipidvalues. Ingestion of soy product has been shown to further improve theeffectiveness of low-fat diets in non-diabetic subjects and the dietarysupplementation in type II diabetic patients may provide an acceptableand effective option for blood lipid control, thereby postponing or evenpreventing drug therapy.

Example 7

Twelve type II diabetic patients (4 female/8 males) with a mean age of65.8±1.6 years, a diabetes duration of 6.0±1.3 years, a mean body massindex of 28.5±1.0, and a mean glycated hemoglobin HbAlc of 7.4±0.4percent were included in the study.

The experiment was an acute, paired, cross-over study in which two testmeals were served during the experiments (A: Standard meal supplementedwith 1 g of stevioside given orally; B: Standard meal given togetherwith 1 g of gelatine (placebo) given orally. The total energy content ofthe test meals was 1725 kJ ( protein 16 E %, fat 30 E %, carbohydrate 54E % ).

Blood samples were drawn from an antecubital vein 30 minutes before and240 minutes after ingestion of the test meal. The arterial bloodpressure was continuously monitored during the experiment. Studentspaired t-test was used for comparing the effects of stevioside withplacebo on the parameters measured. Data are given as mean±SEM.

Stevioside reduced the postprandial blood glucose response by 18±5%(p<0.004) compared to placebo (absolute IAUC 638±55 vs. 522±64mmol/l×240 min; p<0.02) as seen in FIG. 8 a. Stevioside tended tostimulate the insulin response in type II diabetic patients (enhance thearea under the insulin response curve (IAUC)), however the differencedid not reach statistical significance (p=0.09) (FIG. 8 b).

Stevioside significantly reduced the postprandial glucagon levelscompared to placebo (348±46 vs. 281±33; p=0.02) (FIG. 8 c).

Stevioside significantly reduced the postprandial glucagon likepeptide-1 (GLP-1) levels compared to placebo (2208±253 vs. 1529±296;p<0.045) (FIG. 8 d).

Example 8

Four test diets (A: Standard carbohydrate rich laboratory animal diet(Altromin); n=12 (Alt). B: Altromin supplemented with stevioside(Altromin+Stevioside); n=12; (Alt+Ste). C: Soy plus 20% Altromin; n=12;(Soy). D: Soy plus 20% Altromin plus stevioside; n=12; (Soy+Ste)) wereadministered for four weeks to four groups of adult rats. Eachexperimental group consisted of twelve female Goto-Kakizaki with an ageof 9 weeks. The rats received the stevioside (0.025 g/kg bodyweight/day) with the drinking water. By the end of the thirdexperimental week intra-arterial catheters were implanted into thecarotid artery thereby enabling blood sampling during a 240 minutesglucose-tolerance test which was carried out by the end of theexperiment at week 4. Blood samples were drawn after a bolus infusion of2.0 g D-glucose/kg body weight. Plasma concentrations of glucose,insulin, and glucagon were measured during the glucose tolerance test.Immediately before the glucose tolerance test fasting levels oftriglycerides and cholesterol were determined. Concomitantly, thesystolic blood pressure was measured using a tail cuff.

Effects on Plasma-Glucose:

As seen at FIG. 8 and in Table I below stevioside reduced theincremental area (IAUC) under the glucose response curve during theglucose tolerance testing both in the Altromin (p<0.05) and in thesoy+20% Altromin group (Soy) (p<0.001). The relative effect ofstevioside was more pronounced in the group receiving soy+20% Altromingroup compared to the group receiving Altromin. The combination of soyand stevioside synergistically reduced the area under the glucoseresponse curve compared to the Altromin group (p<0.0001) (FIG. 9 a.).

(Plasma glucose was measured using MPR 3, 166 391, Glucose/GOD-PAPMethod from Boehringer Mannheim)

Effects on Plasma Insulin:

The group receiving soy+stevioside (Soy+Ste) has reduced incrementalarea under the insulin response curve compared to theAltromin+stevioside group (Alt+Ste) as seen in FIG. 9 and in Table Ibelow. Considering the concomitant blood glucose responses thisindicates that soy increases the insulin sensitivity. Stevioside did notalter the insulin responses in the Altromin and soy diets when studyingthe total response curve from 0 to 240 minutes. However, in both groupssupplementation of the diets with stevioside significantly improved thefirst phase insulin responses—which is subdued as a characteristicfeature of type II diabetes. The combination of soy+steviosidesynergistically improved the first phase insulin response (p<0.05) (FIG.9 b).

(Plasma insulin was measured using Sensitive Rat Insulin RIA, Cat#SRI-13K from Linco)

Effects on Plasma Glucagon:

Stevioside significantly reduced the area under the plasma-glucagonresponse curve during the glucose tolerance test in both the groupsreceiving Altromin (p<0.003) and soy (p<0.01) (see FIG. 9 c and Table Ibelow).

(Plasma glucagon was measured using Glucagon RIA, Cat #GL-32K fromLinco)

Effects on Blood Pressure:

A marked significant suppression of the systolic blood pressure (p<0.05)(Table I) is elicited by stevioside in combination with either Altromin(Δ=−28 mmHg) or soy (Δ=−21 mmHg) as depicted in FIG. 9 d.

(Blood pressure was measured using TSE Non-Invasive Blood PressureMonitoring System from Technical Scientific Equipment GmbH)

Effects on Body Weight:

The initial weights in the four groups did not differ (FIG. 5).Apparently the combination of soy and stevioside prevented weight gainas seen in FIG. 9 e.

Effects on Triglyceride and Cholesterol:

Stevioside causes a significant suppression of the fasting triglyceridelevels in combination with either Altromin (p<0.05) or soy (p<0.02)(Table I). Soy significantly reduced the fasting triglyceride levelswith or without supplementation of stevioside (p<0.05 and p<0.002,respectively) (Table I). Stevioside given in combination with soysynergistically reduced the fasting total cholesterol levels compared todiets containing Altromin alone (p<0.0001). Soy alone also reduced thetotal cholesterol levels compared to Altromin alone (p<0.002) (FIG. 9 f.and FIG. 9 g) (Table I).

(Plasma cholesterol was measured GOD-PAP from Roche and triglycerideswas measured using GHOD-PAP from Roche)

Stevioside exerts beneficial effects in type II diabetes i.e. reducesblood glucose, suppresses glucagon and improve first phase insulinsecretion. The results also indicates that soy improves insulinsensitivity, a characteristic feature of the metabolic syndrome.Stevioside exerts a pronounced blood pressure reduction both with aswell as without the presence of soy. The combination of stevioside andsoy has a synergistic suppressive effect on blood glucose levels,enhances first phase insulin secretion, suppresses fasting plasmatriglycerides and total cholesterol and the combination of soy andstevioside seems to prevent weight gain. The combination of steviosideand soy appears to possess the potential of an effective treatment of anumber of the characteristic features of the metabolic syndrome i.e.type II diabetes, hypertension, dyslipidemia and obesity. TABLE I IAUCIAUC IAUC p-insulin p-insulin IAUC Change in blood p-glucose (ng/ml ×240 (ng/ml × 30 p-glucagon (pg/ pressure (mmHg) TriglyceridesCholesterol Group (mM × 240 min) min) min) ml × 240 min) From week 0 to4 (mM) (mM) Altromin 991 ± 96 317 ± 55 11 ± 4 21918 ± 1467 5 ± 4 0.72 ±0.10 2.51 ± 007  Altromin + 757 ± 53 375 ± 42 19 ± 4 17023 ± 1449 −23 ±6  0.50 ± 0.04 2.28 ± 0.18 Stevioside Soy + 20% Altromin 820 ± 75 218 ±22  9 ± 2 26200 ± 2410 8 ± 3 0.49 ± 0.04 2.13 ± 0.08 Soy + 20%Altromin + 439 ± 56 248 ± 27 24 ± 5 17229 ± 1819 −13 ± 5  0.37 ± 0.021.84 ± 0.06 SteviosideTable I: Areas under the p-glucose, -insulin and -glucagon responsecurves during the glucose tolerance test in the four experimentalgroups. Change in systolic blood pressure at start and at end of thestudy period. Fasting plasma-triglyceride and -total cholesterolconcentrations by the end of the study.

1. A method of treating medical conditions in a mammal resulting fromelevated plasma glucose which comprises administering to a mammal inneed of such treatment a therapeutically effective amount of a bicyclo[3.2.1]octan in a double ring system having a basic chemical skeletal ofa kaurene structure having the structural formula II:

or an analogue, derivative or metabolite thereof, wherein the substanceresponds in the mammal only at plasma glucose concentrations that areelevated above normal levels.
 2. The method according to claim 1,wherein the condition is non-insulin dependent diabetes mellitus,metabolic syndrome, hypertension, or appetite.
 3. The method accordingto claim 1, wherein the response of the bicyclo [3.2.1]octan isinitiated by a plasma glucose concentration of 6 mmol/l or greater. 4.The method according to claim 1, wherein the response of the bicyclo[3.2.1]octan is initiated by a plasma glucose concentration of 6 mmol/lor greater.
 5. The method according to claim 1, wherein the bicyclo[3.2.1]octan is selected from the group consisting of steviol,isosteviol, glucosilsteviol, gymnemic acid, steviolbioside, steviosidRebaudioside A, Rebaudioside B, Rebaudioside C, Rebaudioside D,Rebaudioside E, Dulcoside A, their pharmaceutically acceptable analoguesand their pharmaceutically acceptable derivates.
 6. The method accordingto claim 1, wherein the bicyclo [3.2.1]octan is isolated from a plantsource.
 7. The method according to claim 1, wherein the bicyclo[3.2.1]octan is administered in combination with at least one soyprotein or in combination with at least one soy protein and at least oneisoflavone.
 8. The method according to claim 1, wherein the bicyclo[3.2.1]octan is present a composition that includes a carrier.
 9. Themethod according to claim 1, wherein the bicyclo [3.2.1]octan isadministered orally to the mammal and is self-regulating.
 10. The methodaccording to claim 1, wherein the bicyclo [3.2.1]octan is administeredin an amount effective to treat non-insulin dependent diabetes mellitusin the mammal.
 11. The method according to claim 1, wherein the bicyclo[3.2.1]octan is administered in an amount effective to treat metabolicsyndrome in the mammal.
 12. The method according to claim 1, wherein thebicyclo [3.2.1]octan is administered in an amount effective to stimulateinsulin production.
 13. The method according to claim 1, wherein thebicyclo [3.2.1]octan is administered in an amount effective to reduceblood glucagon concentrations in the mammal.
 14. The method according toclaim 1, wherein the bicyclo [3.2.1]octan is administered in an amounteffective to treat hypertension in the mammal.
 15. The method accordingto claim 1, wherein the bicyclo [3.2.1]octan is administered in anamount effective to suppress fasting plasma triglycerides in the mammal.16. The method according to claim 1, wherein the bicyclo [3.2.1]octan isadministered in an amount effective to suppress total cholesterol levelsin the mammal.
 17. The method according to claim 1, wherein the bicyclo[3.2.1]octan is administered in an amount effective to act as a dietarysupplement.
 18. The method according to claim 1, wherein the bicyclo[3.2.1]octan is administered in an amount effective to act as anappetite suppressant in the mammal.
 19. The method according to claim 1,wherein the bicyclo [3.2.1]octan is steviol, isosteviol,glucosilsteviol, gymnemic acid, steviolbioside, stevioside, RebaudiosideA, Rebaudioside B, Rebaudioside C, Rebaudioside D, Rebaudioside E orDulcoside A.